IL-13在哮喘中的作用

支气管哮喘是以Th2型细胞因子增高和气道高反应性为特征的变态反应性疾病。白细胞介素 13(IL 13)是近年来新克隆的Th2型细胞因子 ,它通过诱导B细胞增殖和分化 ,促进IgE合成 ,活化嗜酸性粒细胞 ,延长嗜酸性粒细胞的存活 ,诱导气道高反应性等机制 ,在哮喘的发生中起重要的作用。     

070 IL-13在哮喘中的作用

支气管哮喘是以Th2型细胞因子增高和气道高反应性为特征的变态反应性疾病.白细胞介素-13(IL-13)是近年来新克隆的Th2型细胞因子,它通过诱导B细胞增殖和分化,促进IgE合成,活化嗜酸性粒细胞,延长嗜酸性粒细胞的存活,诱导气道高反应性等机制,在哮喘的发生中起重要的作用.     

白介素13在哮喘发病机制中的作用

Th2淋巴细胞的优势应答及其释放的细胞因子被认为在哮喘的发病过程中起关键性作用。IL-13可不依赖于其它Th2型细胞因子及嗜酸粒细胞、Ig-E介导的途径而单独诱发哮喘的所有症状。IL-13基因多态性与哮喘的易感性也密切相关。其拮抗剂的应用有望成为哮喘病防治的新方法。     

IL-13与支气管哮喘

支气管哮喘是由多种细胞特别是肥大细胞、嗜酸性粒细胞和T淋巴细胞参与的慢性气道炎症。哮喘的发生过程中有多种细胞因子参与 ,白介素 13(IL 13)是近年来新克隆的淋巴因子。IL 13作为Th2型细胞因子成员 ,具有多种生物学功能。通过激活嗜酸性粒细胞 ,减少其凋亡 ,促进IgE分泌等机制 ,参与哮喘炎症的维持 ,诱导气道高反应性及小气道结构重建     

IL-13对成纤维细胞IL-13受体和IL-4受体表达的影响

目的研究IL-13对人肺成纤维细胞株HFL-1和人肝星状细胞株LX-2 IL-13受体和IL-4受体表达的调节作用。方法 RT-PCR法检测HFL-1细胞株和LX-2细胞株IL-13Rα1、IL-4R和IL-13Rα2 mRNA的表达;凝胶电泳定量软件Image Tool2.0对RT-PCR电泳条带进行光密度分析;ELISA法检测HFL-1细胞株和LX-2细胞株分泌可溶型IL-13Rα2以及检测细胞裂解液总IL-13Rα1、IL-4R和IL-13Rα2含量。结果 IL-13(5～100 ng/ml)对HFL-1细胞株和LX-2细胞株表达IL-13Rα1和IL-4R无影响;IL-13为5、10、20 ng/ml时能诱导HFL-1细胞株表达IL-13Rα2并呈现剂量依赖,当IL-13为50 ng/ml时,对HFL-1细胞株IL-13Rα2表达的诱导作用明显减弱,IL-13 100 ng/ml组没有检测到HFL-1细胞株IL-13Rα2的表达;LX-2细胞株IL-13Rα2表达缺失且IL-13不能诱导LX-2细胞株表达IL-13Rα2。结论 IL-13不能上调人肺成纤维细胞株HFL-1和人肝星状细胞株LX-2表达功能型受体IL-13Rα1和IL-4R,表明IL-13不能通过上调IL-13Rα1和IL-4R表达量来放大自身作用;一定浓度的IL-13能诱导人肺成纤维细胞株HFL-1表达抑制型受体IL-13Rα2,表明IL-13的自身负调控。     

IL-4、IL-13蛋白表达及抗IL-4、IL-13人源单链抗体的筛选

目的:表达IL-4和IL-13蛋白,从人源单链抗体文库中分别筛选抗IL-4和抗IL-13单链抗体.方法:采用RT-PCR从健康志愿者外周血单核细胞(PBMC) mRNA中扩增IL-4和IL-13 cDNA;构建硫氧还蛋白融合表达载体,转化大肠杆菌BL21,IPTG诱导表达并对表达产物进行纯化鉴定.以生物素化的IL-4和IL-13为抗原从前期构建的人源抗体文库中采用噬菌体展示技术分别筛选抗IL-4和抗IL-13人源单链抗体(scFv).结果:扩增的IL-4 cDNA大小为280 bp,表达的融合蛋白大小为27 kD左右.扩增的IL-13 cDNA大小为252 bp,表达的融合蛋白大小为25 kD左右.分别以生物素化的IL-4和IL-13蛋白为抗原,采用噬菌体展示技术对人源抗体文库进行3轮富集后,分别有大约37％的scFvs与IL-4有结合特性,有约27％的scFvs与IL-13有结合特性.筛选了4株分别与IL-4和IL-13结合能力强的单链抗体进行了Westem blot鉴定和测序.结论:成功筛选到抗IL-4和抗IL-13人源性单链抗体.     

IL-4R、IL-13、ADAM33基因多态性与中国皖南地区汉族人群支气管哮喘的相关性研究

目的:探讨IL-4受体基因Arg551Gln(rs1801275)、IL-13基因Arg130Gln(rs20541)、ADAM33基因T1(rs2280091)位点基因多态性与中国皖南地区汉族人群支气管哮喘的相关性。方法:采用病例-对照的方法,用聚合酶链反应及直接基因测序法比较116例支气管哮喘组与70例正常人对照组之间基因型、等位基因频率的差异。结果:哮喘组和对照组IL-4受体基因Arg551Gln位点和IL-13基因Arg130Gln位点的基因型和等位基因型频率的差异有统计学意义,ADAM33基因T1位点基因型哮喘组和对照组差异有统计学意义,等位基因型频率在哮喘组和对照组差异无统计学意义。结论:提示IL-4R Arg551Gln(rs1801275)位和IL-13基因Arg130Gln(rs20541)位的多态性可能与中国皖南地区汉族哮喘有相关性;ADAM33基因(rs2280091)T1位点位的多态性可能与中国皖南地区汉族哮喘无相关性。     

青霉素过敏反应与IL-4、IL-13、IFN-γ

目的 :探讨青霉素类抗生素过敏与IL 4、IL 1 3、IFN γ之间的关系。方法 :采用ELISA法检测了 1 45例过敏病人和 62例正常人血清中的IL 4、IL 1 3和IFN γ浓度 ;采用RAST法检测了过敏病人和正常人血清中的 8种特异性IgE抗体 (BPO PLL、PVO PLL、APO PLL、AXO PLL、BPA PLL、PVA PLL、APA PLL、AXA PLL)。结果 :特异性IgE抗体阳性的青霉素过敏病人组IL 4、IL 1 3、IFN γ血清浓度显著高于正常组 (P <0 .0 1 ) ;特异性IgE抗体阴性的青霉素过敏病人组IL 4、IFN γ浓度显著低于正常组 (P <0 .0 1 ) ;IL 4…     

IL-4和IL-4受体基因多态性与成人变应性哮喘的关系

目的　研究白细胞介素 4 (IL 4 )、IL 4受体α链的 2个基因多态性位点与中国成人变应性哮喘的关系。方法　采用病例对照方法 ,用聚合酶链反应 限制性片段长度多态性方法 (PCR RFLP)对IL 4启动子区C - 5 89T和IL 4Rα链Q5 76R进行基因分型。结果　IL 4C - 5 89T与中国成人变应性哮喘无关 ,然而 ,变应性哮喘组IL 4Rα链 5 76R R频率显著性高于对照组 (χ2 =9.36 9,P <0 .0 1;OR =3.797) ,且与血浆高IgE相关。结论　IL 4Rα链 5 76R R基因型是中国成人变应性哮喘的基因危险因子     

IL-13基因与哮喘患者关联性的研究进展和展望

哮喘属于气道慢性炎症性疾患,其发生是遗传因素与环境因素相互作用的结果。哮喘的遗传率约为36%-79%。国内外对哮喘家系或人群的连锁分析、关联研究等分子遗传学研究显示,哮喘相关基因及其多态性至少与人类多条染色体上的100多个基因关联。由于哮喘呈多基因、复杂性遗传方式,涉及的基因及单核苷酸多态性(SNP)位点众多,国内外对相关基因的研究结果多有不一,使得哮喘发病的分子机制至今不详。位于人类染色体5q31上的IL-13基因可能在哮喘的发病机制中具有重要作用。外显子组捕获测序以及即将面世的第三代测序或单分子实时测序技术,是目前寻找哮喘易感基因的最好方法。     

The role of IL-13 in IgE synthesis by allergic asthma patients

IgE antibodies play a crucial role in allergic type I reactions. Only IL-4 and IL-13 are able to induce an immunoglobulin isotype switch to IgE in B cells. A major question is to what extent these cytokines contribute to the production of IgE in allergic patients. To address this question we used an in vitro culture system in which the production of IgE is dependent on endogenously produced IL-4 and IL-13. In cultures of purified T and B cells from allergic asthma patients and non-atopic controls, T cells were polyclonally stimulated to obtain IL-4, IL-13 and subsequently IgE secretion. The absolute amount of IgE produced was not significantly different between patients and controls. When neutralizing IL-4 antibodies were included during culture, the production of IgE was dramatically inhibited in both patients and controls (production of IgE was reduced to 12%). However, neutralization of IL-13 led to a significantly stronger inhibition of IgE production in the patient group: production of IgE was reduced to 23 ± 3% versus 50 ± 10% in the control group. Corresponding with these results, we also observed a higher production of IL-13 by the patients, while the production of IL-4 was not significantly different. A more detailed analysis of the production of IL-13 revealed that patients' T cells were less sensitive to a negative signal controlling IL-13 production. Our results indicate that, at least in vitro, IgE production in allergic asthma patients is more dependent on IL-13 than in non-atopics, due to enhanced IL-13 production and to enhanced IgE production in response to IL-13.     

Th2 Cytokines IL-4 and IL-13 Downregulate Paxillin Expression in Bronchial Airway Epithelial Cells

Asthma is characterized by infiltration and shedding of the bronchial epithelium. The Th2 cytokines IL-4 and IL-13 are involved in the cellular recruitment and infiltration seen in asthma. The effects of IL-4 and IL-13 on cell-matrix interactions and epithelial shedding are unknown. We hypothesize that bronchial airway epithelial cells (BAEC) express paxillin, a structural focal adhesion protein, and downregulation of paxillin by Th2 cytokines lead to BAEC hyperpermeability. We showed by confocal microscopy the presence of paxillin in BAEC. We demonstrated by Western blot analysis that IL-4 and IL-13 stimulation results in downregulation of paxillin production. IL-4 and IL-13 stimulation decreased epithelial cell-matrix attachment as measured by electrical cell-substrate impedance sensing system (ECIS). Our results suggest that Th2 cytokines IL-4 and IL-13 downregulate paxillin production by BAEC, thereby disrupting the cell-matrix interactions. This may help explain the epithelial shedding and epithelial membrane hyperpermeability that occurs in asthma.     

Therapeutic targeting of IL-4-and IL-13-responsive cells in pulmonary fibrosis

Severe forms of idiopathic interstitial pneumonia (IIP), such as usual interstitial pneumonia (UIP), can be impervious to modern steroid and immunosuppressive treatment regimens, thereby emphasizing the need for novel effective therapies. Understanding the cytokine networks that may affect immune and structural cell activation and, hence, the progression of these fatal fibrotic diseases, has been a focus in our research. In this regard, we have examined the role of interleukin (IL)-4 and IL-13 and their respective receptor subunits in this process. Examination of clinical surgical lung biopsies (SLBs) showed that IIP is characterized by the abnormal, heightened expression of the receptor subunits that bind IL-4 and IL-13. Specifically, IL-4Rα and IL-13Rα2 (the high-affinity IL-13 receptor subunit) was present in greater abundance in SLBs and fibroblasts from IIP patients compared with normal patients, who exhibited no evidence of pulmonary fibrosis. These clinical findings prompted us to investigate whether the targeting of pulmonary cell types that were highly responsive to IL-4 and IL-13 was a viable therapeutic option in IIP. Using a chimeric protein comprised of human IL-13 and a truncated version of an exotoxin from Pseudomonas (abbreviated IL13-PE), we observed that IL13-PE selectively targeted human pulmonary fibroblasts grown from IIP SLBs, whereas it had a minimal effect on fibroblasts grown from biopsies from normal patients. In murine models characterized by abnormal airway or interstitial fibrotic responses, the intranasal administration of IL13-PE significantly attenuated the fibrotic response through the targeting of IL-4Rα-and IL-13Rα2-expressing pulmonary cells, including monocytes, macrophages, and pulmonary fibroblasts. Together, these data demonstrate that IL-4 and IL-13 are required for the initiation and maintenance of pulmonary fibrosis, and highlight the importance of further investigation of anti-fibrotic therapeutics that prevent the action of both cytokines during clinical pulmonary fibrosis.     

Inhaled vs subcutaneous effects of a dual IL-4/IL-13 antagonist in a monkey model of asthma

Background:  Pitrakinra is a recombinant protein derived from human interleukin-4 (IL-4) that binds to IL-4Rα and acts as a competitive antagonist of IL-4 and IL-13. The studies reported here compare the dose-ranging effects of pitrakinra on allergen-induced airway hyperresponsiveness (AHR) and airway eosinophilia when administered subcutaneously (s.c.) or by inhalation to the Ascaris suum -sensitive cynomolgus monkey for the purpose of elucidating the primary site of pitrakinra's anti-asthmatic action.
Methods:  Airway responsiveness to inhaled methacholine and bronchoalveolar lavage cell composition was determined before and after three allergen exposures with a 1-week course of twice-daily (b.i.d.) s.c. or inhaled pitrakinra or placebo treatment.
Results:  Treatment with s.c. pitrakinra significantly reduced allergen-induced AHR, with a maximum effect of a 2.8- to 3.8-fold increase in methacholine PC₁₀₀ relative to control ( P  < 0.05) observed at b.i.d. s.c. doses of 0.05–0.5 mg/kg. Inhaled pitrakinra also significantly reduced AHR with a similar maximum effect of a 2.8- to 3.2-fold increase in methacholine PC₁₀₀ relative to control ( P  < 0.05) at nominal b.i.d. doses of 3–100 mg. The maximal effect on AHR following inhalation was observed at a plasma concentration which exhibited no efficacy via the subcutaneous route. The effect of pitrakinra on lung eosinophilia was not statistically significant following either route of administration, although lung eosinophil count was reduced in all studies relative to control.
Conclusion:  Local administration of pitrakinra to the lung is sufficient to inhibit AHR, one of the cardinal features of asthma, indicating the therapeutic potential of inhaled pitrakinra in the treatment of atopic asthma.     

IL-13和CD86对9HTE细胞株和哮喘患儿PBMC表达CD86水平的影响

目的 了解IL-13和CD86在哮喘发病中的作用及anti-CD86 McAb治疗哮喘的机理。方法 随机选择门诊哮喘急性发作期患儿30例，正常健康儿童30例作为健康对照组。分别用rhIL-13、TNF-α和rhIL-13 TNF-α干预9HTE人气道上皮细胞株和哮喘患儿外周血单个核细胞(peripheral blood mononuclear cell，PBMC)，用直接免疫荧光流式细胞术(FCM)检测9HTE人气道上皮细胞株表达CD86的百分率、PBMC中CD14^*细胞表达CD86的平均荧光强度(MFI)及脂多糖(LPS)刺激后PBMC中CD19^ CD86^ 双阳性细胞百分率。结果 (1)分别用TNF-α、rhIL-13和TNF-α rhIL-13干预9HTE细胞株后CD86的表达，三组之间表达的百分率差异无统计学意义；rhIL-13 anti-CD86 McAb干预9HTE细胞株后，9HTE细胞表达CD86水平较其他组降低，差异有统计学意义。(2)哮喘组中的空白组、IL-13组、TNF-α组、TNF-α IL-13组及anti-CD86组的CD14^ CD86^ 的平均荧光强度和CD19^ CD86^ 百分率均较相应的对照组高，差异有统计学意义。(3)TNF-α IL-13干预后，CD14^ CD86^ 的平均荧光强度和CD19^ CD86^ 百分率较对应的空白组、IL-13组和TNF-α组明显增高，差异有统计学意义；(4)anti-CD86 McAb干预后，CD14^ CD86^ 的平均荧光强度和CD19^ CD86^ 百分率较对应的其他组明显减低，差异有统计学意义。结论 IL-13可诱导9HTE细胞株、PBMC中的CD14^ 细胞和CD19^ 细胞表达CD86；anti-CD86 McAb可阻断9HTE细胞株、PBMC中的CD14^ 细胞和CD19^ 细胞表达CD86。提示IL-13和CD86在哮喘发病中起重要作用，anti-CD86 McAb治疗哮喘具有一定的可行性。     

Immunoregulatory properties of IL-13 in patients with inflammatory bowel disease; comparison with IL-4 and IL-10

Activated monocytes with increased expression of proinflammatory cytokines play a major role in inflammatory bowel disease (IBD). Immunoregulatory cytokines such as IL-4 and IL-10 can effectively suppress the proinflammatory response of activated monocytes. IL-13 is a recently described antiinflammatory agent in vitro. The aim of our study was to determine the in vitro immunosuppressive capacity of IL-13, IL-4 and IL-10 in patients with IBD. Peripheral blood monocytes were isolated from 27 patients with ulcerative colitis (UC), 27 patients with Crohn's disease (CD) and 16 healthy controls. Cells were stimulated with pokeweed mitogen (PWM) after treatment with IL-13, IL-4 and IL-10, and secretion of IL-1β, tumour necrosis factor-alpha (TNF-α) and IL-6 was assessed using sandwich ELISA systems. Peripheral blood monocytes secreted significantly increased amounts of TNF-α and IL-6 under stimulation with PWM in patients with CD, while UC patients showed significantly elevated levels of IL-1β. The antiinflammatory cytokines IL-13, IL-4 and IL-10 were all capable of inhibiting monocyte secretion of IL-1β in a dose-dependent manner. With regard to IL-13 and IL-4, there was no significant suppression of TNF-α and IL-6 in patients with active IBD. By contrast, IL-10 was able to down-regulate all proinflammatory cytokines in active IBD as well as in controls. Proinflammatory cytokines from patients with inactive IBD could be significantly down-regulated by all three immunoregulatory cytokines. The inhibitory effect of IL-13 on TNF-α and IL-6 production in differentiated macrophages was diminished in IBD patients, as well as in controls. In disease controls we also observed a reduced inhibition of TNF-α and IL-6 after treatment with IL-13. In conclusion, the antiinflammatory activity of IL-13 is partially reduced in patients with active IBD. The hyporesponsiveness of activated and differentiated monocytes to IL-13 and IL-4 does not seem to be a disease-specific phenomenon.     

哮喘患者IL-13基因多态性与IL-13、TIgE水平相关性研究

目的:探讨白细胞介素13(IL-13)基因内含子区+1923C/T多态性与哮喘患者外周血单个核细胞(PBMC)产IL-13、血浆总IgE(TIgE)水平及其相关性。方法:用聚合酶链反应和限制性片段长度多态性(PCR/RFLP)方法检测哮喘组与对照组+1923C/T位点多态性。IL-13、血浆总IgE采用ELISA法。结果:+1923位点等位基因C、T频率在两组间分布的差异具有显著性(X2=9.30,P<0.01);等位基因T与哮喘关联,OR(T/C)=1.87,95%CI=1.25-2.80,P<0.01。两组基因型(TT、CT、CC)频率的分布差异亦有显著意义(X2=9.92,P<0.01)。其优势比:OR(TT/CC)=3.76,95%CI=1.52-9.29,P<0.01;OR(CT/CC)=2.10,95%CI=1.11-3.95,P<0.05;OR(TT/CT)=1.79,95%CI=0.77-4.19,P>0.05。哮喘组中TT、TC基因型人群PBMC产IL-13及TIgE水平与同组及对照组CC基因相比较差异均有显著性(P<0.01)。结论:IL-13基因+1923位点多态性是影响哮喘的重要候选基因,T等位基因与哮喘关联。